Hydrogen-Deuterium eXchange coupled to Mass Spectrometry (HDX-MS) analysis to investigate the solvent accessibility of the full human 20S proteasome alone or in complex with PA28 regulatory particles. Our work reveals a reciprocal crosstalk between the inner and outer rings of the human 20S proteasome. HDX-MS experiments were performed on a Synapt-G2Si (Waters Scientific, Manchester, UK) coupled to a Twin HTS PAL dispensing and labelling robot (LEAP Technologies, Carborro, NC, USA) via a NanoAcquity system with HDX technology (Waters, Manchester, UK).